Chromatin architecture plays major roles in gene regulation as well as in the repair of DNA damaged by endogenous or exogenous factors, such as after radiation. Opening up the chromatin might provide the necessary accessibility for the recruitment and binding of repair factors, thus facilitating timely and correct repair. The observed formation of ionizing radiation-induced foci (IRIF) of factors, such as 53BP1, upon induction of DNA double-strand breaks have been recently linked to local chromatin decompaction. Using correlative light and electron microscopy (CLEM) in combination with DNA-specific contrasting for transmission electron microscopy or tomography, we are able to show that at the ultrastructural level, these DNA damage domains reveal a chromatin compaction and organization not distinguishable from regular euchromatin upon irradiation with carbon or iron ions. Low Density Areas (LDAs) at sites of particle-induced DNA damage, as observed after unspecific uranyl acetate (UA)-staining, are thus unlikely to represent pure chromatin decompaction. RNA-specific terbium-citrate (Tb) staining suggests rather a reduced RNA density contributing to the LDA phenotype. Our observations are discussed in the view of liquid-like phase separation as one of the mechanisms of regulating DNA repair.
Keywords: CLEM; ChromEMT; DNA and RNA cytochemistry; DNA repair; DNA-specific staining; Osmium ammine B; carbon ions; chromatin structure; electron microscopy; radiation-induced damage.